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Three-dimensional correlative light and electron microscopy (3D CLEM) is attaining popularity as a potential technique to explore the functional aspects of a cell together with high-resolution ultrastructural details across the cell volume. To perform such a 3D CLEM experiment, there is an imperative requirement for multi-modal probes that are both fluorescent and electron-dense. These multi-modal probes will serve as landmarks in matching up the large full cell volume datasets acquired by different imaging modalities. Fluorescent nanodiamonds (FNDs) are a unique nanosized, fluorescent, and electron-dense material from the nanocarbon family. We hereby propose a novel and straightforward method for executing 3D CLEM using FNDs as multi-modal landmarks. We demonstrate that FND is biocompatible and is easily identified both in living cell fluorescence imaging and in serial block-face scanning electron microscopy (SB-EM). We illustrate the method by registering multi-modal datasets. [ABSTRACT FROM AUTHOR]

Quantifying the density and locating the position of antigens on cell surface has been a challenge in molecular biology research. The challenge lies in the need for a chemically and photophysically stable fluorophore to achieve the required sensitivity and accuracy. Here, we present a method suitable for the purpose by using lipid-encapsulated fluorescent nanodiamonds (FNDs) of 35 nm in diameter as biolabels. The encapsulation of FNDs in biotinylated phospholipids not only facilitates good dispersion of the particles in biological buffers, but also endows them with high specific targeting ability. We demonstrated a viable application of the technique for biotin-mediated immunostaining of antigens on fixed human cells, identifying their positions by two-color confocal fluorescence imaging, and determining their densities by magnetically modulated fluorescence detection. A binding capacity of 6 ± 1 × 104 antigens/cell was measured specifically for CD44 on HeLa cell surface. The result agreed well with the assay of R-phycoerythrin-conjugated antibodies by flow cytometry, supporting the reliability of this new nanoparticle-based method. [ABSTRACT FROM AUTHOR]